Our research has been concerned with the expression of regulatory mechanisms, in intact cardiac preparations, of automatic neurotransmitters and other hormones and drugs that act through cyclic AMP and protein phosphorylation. This messenger and its mode of action appears to lack specificity, i.e. many agents elevate cyclic AMP in heart; the variety of substrates for protein kinase is large. One mechanism of specificity is suggested by our finding that in contrast to catecholamines, prostaglandin E1 stimulates cyclic AMP production and protein kinase but with no evidence of protein phosphorylation. Catecholamines produce protein phosphorylation (including the contraction-regulating protein, troponin) and this action always associated with solubilization of the activated protein kinase. This step appears to be the critical one that is absent in the response to PGE1. Another mechanism of autonomic neurotransmitter regulation was discovered from the observation that depolarization of cardiac muscle blocked the entire sequence of reactions following catecholamine-receptor interaction: cyclic AMP-protein kinase-phosphorylase pathway, and increased contractility. This is due to stimulation of acetylcholine release from parasympathetic nerve endings which either antagonizes binding of catecholamine to receptors or causes the formation of an antagonist second messenger, presumably cyclic GMP. The site of muscarinic action appears to involve the mechanism of coupling of the hormone receptor complex to adenylate cyclase.